The goal of this application is to identify biomarkers of oral squamous cell carcinoma (OSCC) initiation and progression to metastasis by integrating experimental and computational analysis approaches. Specifically, this study will investigate the role of a homeostatic cellular network consisting of the metabolic pathway of protein N-glycosylation, the Wnt/?-catenin and the Hippo/YAP signaling pathways in driving pro-tumorigenic activities in oral epithelia leading to the development and progression of OSCC. The elucidation of this network's key regulators and their downstream effectors offers the possibility to identify novel druggable targets in the OSCC pathogenic pathways and thus to advance the development of more effective and lasting OSCC therapies. Previous studies provide evidence that the interactions of this network are controlled by the N-glycosylation-regulating gene, DPAGT1, the Wnt pathway effector, ?-catenin, and the Hippo pathway effector, YAP. The three factors are co-dependent in their expression and activity, and aberrant activation of the DPAGT1/?-catenin/YAP (glyco-?Y) network promotes cancerous phenotypes. Of significance, the interrogation of genome-wide oral cancer datasets highlights increased activity of DPAGT1, ?-catenin and YAP as a prominent feature of human OSCC. Activation of the glyco-?Y signaling network is hypothesized to drive OSCC by promoting oral epithelial cell proliferation, survival and migration. Here, this hypothesis will be tested through the: (1) identification of gene targets significantly deregulated upon transcriptional inhibition of each of these effectors in OSCC cell lines; (2) definition of pro-tumorigenic gene expression signatures at different stages of primary human OSCC development, and elucidation of their role in driving OSCC to metastasis; (3) development of a novel computational method to associate genetic and epigenetic factors, which include somatic mutations and copy number alterations, DNA methylation and microRNAs, with gene expression signatures of OSCC pathogenic pathway activity. The experimental strategies will utilize a panel of human OSCC cell lines, human premalignant tissues, and human OSCC specimens. Unbiased global RNA-sequencing (RNASeq) interrogation using treated OSCC cell lines, and human tissue specimens at different stages of OSCC development will identify downstream effectors of the glyco-?Y network, and will point to novel candidate targets which will be validated both experimentally, as well as computationally by interrogating large multi-omic oral cancer datasets. The deliverables of this application include: (i) validation of glyco-?Y network involvement in OSCC development; (ii) identification of novel gene expression markers of human OSCC initiation and progression; (iii) development of a new integrative genomics tool that can identify candidate genetic and epigenetic drivers of signature-defined pathway activity in OSCC.